Bacterial Toxin Research Citations

2.1. Chondrocyte isolation and culture

… Samples were placed in sterile saline (0.9% NaCl) solution with gentamicin sulfate (50 mg/l) and amphotericin B (250 μg/ml). The cartilage samples were transported chilled (approx. 5 °C) to the laboratory. Isolation and expansion of chondrocytes were performed as previously described (Ley et al., 2011). Briefly, the chondrocytes were expanded to passage 1 and then seeded at 20,000 cells/cm2 in chondrogenic medium to maintain the phenotype. On day 4, cells were stimulated with LPS (10 ng/ml, Escherichia coli 055:B5; List Biological Laboratories, Campbell, CA, USA) or kept untreated (controls) for 24 h. Cells were grown to confluence and harvested on day 5 and immediately frozen and stored at −80 °C until further analyses.

• Product #203A – LPS from Escherichia coli O55:B5

… goat anti-CTB (1:2k, List Biological Laboratories, catalog # 703, RRID: AB_10013220), …

• Product #703 – Anti-Cholera Toxin B Subunit (Goat)

4.6. Serological analysis of B. pertussis specific antibodies

On the day of processing, a cardiac puncture was performed on each mouse. Approximately 1 mL of blood was removed from each mouse. The samples were centrifuged for 2 min at 14,000 × g. Serum was collected and stored at − 80 °C until analyses were performed. Serological responses specific to B. pertussis antigens were quantified by ELISA. High-binding microtiter plates were coated with PT (50 ng/well) (PT#180, LIST Biologicals), FHA (50 ng/well) (Enzo Life Sciences), or PRN (50 ng/well) (PRN#187, LIST Biologicals) as described in Boehm et al. [52]. …

• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #187 – Pertactin from B. pertussis (69 kDa Protein)

… There followed an incubation in anti-CT-B made in goat (List Biological #703; 1:4000 dilution in TBS-TX) and 3% horse serum for 2-3 days at 4°C …

• Product #703 – Anti-Cholera Toxin B Subunit (Goat)

… After EAE induction, intraperitoneal injections of 200 ng pertussis toxin (List Biological Laboratories, USA) in the first hour and 400 ng pertussis toxin in 200 µl PBS were administered at the 48th hour. …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

Colony-forming assay

Colony-forming assay (CFA) was performed as previously reported (Sasamoto et al., 2020). Briefly, limbal epithelial cells were seeded on the mitomycin C (MMC) (MilliporeSigma)-treated 3T3-J2 feeder cell layer at 500 cells per well on 6-well plates. The cells were cultured in keratinocyte culture medium (KCM) supplemented with 10 ng/mL KGF and 10 μM Y-27632. KCM was composed of DMEM without glutamine and Ham’s F-12 Nutrient Mix (Thermo Fisher Scientific) combined at 3:1 ratio, supplemented with 10% FBS, 0.4μg/ml hydrocortisone hydrogen succinate (MilliporeSigma), 2nM 3,3′,5-triiodo-l-thyronine sodium salt (MilliporeSigma), 1 nM cholera toxin (List Biological Laboratories, Campbell, CA), …

• Product #100B – Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

Experimental Human Endotoxemia (IV LPS Administration)

All experimental endotoxemia-related procedures were performed as described previously and were identical on both LPS challenge days (days 0 and 7) [19, 22]. In short, subjects were admitted to the research unit of the Radboud University Medical Center for 8 h. Subjects had to refrain from alcohol and caffeine (24 h) and food and drinks (12 h) prior to LPS administration. A radial artery catheter (BD Infusion Therapy Systems, Sandy, UT, USA) and antebrachial venous cannula were placed to allow serial blood sampling, hemodynamic monitoring, and administration of fluids and LPS, respectively. In the 45 min prior to LPS administration, hydration fluids (2.5% glucose/0.45% sodium chloride) were administered as a 1.5-L prehydration bolus to reduce the risk of vasovagal collapse [23] and thereafter at a rate of 150 mL/h for the remainder of the experiment. Directly after prehydration, a bodyweight-adjusted bolus dose of 1 ng/kg LPS (Escherichia coli-derived, Type O113, lot No. 94332B1; List Biological Laboratories, Campbell, CA, USA) was administered. Blood samples were serially obtained to construct time-concentration curves of circulating cytokines.

• Product #9433 – GMP LPS Lipopolysaccharide List™ Hpt™ From Escherichia Coli Type O113

… Bordetella Pertussis toxin List Biological Labs Cat. #: 180 …

MOG35-55-induced EAE
EAE was induced in mice using MOG35-55 as described by Bittner et al. (2014). Briefly, 8-week-old endothelial-specific Arf6 knockout mice (Arf6f/f; Cdh5-CreERT2) and their control sibling littermates or C57BL/6J mice were immunized with MOG35-55 in CFA emulsion followed by administration of pertussis toxin (PTX) in PBS at day 0 and again at day 2. …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

2.7.4 Tracer intramuscular injection

Adult female Lister Hooded rats (220–290 g; Charles River; husbandry conditions as above), either naïve (n = 5) or 10 weeks after SCI (spinal C7 bilateral contusion, as above; n = 3) had the right biceps brachii intramuscularly injected with CTB. Rats were anesthetized with isoflurane (5% induction and 2.5% maintenance in 1 L/min O2). Analgesia (carprofen, CarprieveTM, 5 mg/kg) was administered subcutaneously, and the right forelimb was shaved and disinfected with 4% w/v chlorhexidine gluconate. The skin was incised and retracted, and the fascia was carefully opened for clear visualization of the two heads of the muscle. A total of 12 μl of 0.5% CTB diluted in distilled water (104 Cholera Toxin B Subunit; #10433A1, List Biological Laboratories, Inc.) was injected into the right biceps using 10 μl Hamilton syringes (#80300, 26s-gauge, Hamilton). The short head of the biceps received 8 μl of CTB, divided into two deep injections of 2 μl and two superficial injections of 1 μl aiming the motor endplates (Tosolini & Morris, 2012) and two deep injections of 1 μl aiming at the region halfway between the proximal end of the muscle and the motor endplates (Figure 6a). The long head of the biceps received 4 μl of CTB, divided into one deep injection of 2 μl and one superficial injection of 1 μl aiming the motor endplates, and one deep injection of 1 μl aiming the region halfway between the proximal end of the muscle and the motor endplates (Figure 6a). The muscle was wiped before and after injections with a cotton bud. The needle was inserted parallel to the muscle fibers, the tracer was injected slowly, and the needle was kept in place after injection for at least 30 s. After injections, the skin was sutured, and saline administered subcutaneously. Rats recovered from anesthesia in a heated recovery chamber.

• Product #104 – Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt

Surgical procedures

Mice were anesthetized with an intraperitoneal injection of the ketamine and xylazine mixture (100 mg/kg and 10mg/kg, respectively). For the creation of SNI, muscles were displaced to expose the right sciatic nerve, and the nerve was ligated proximal to its trifurcation. The sciatic nerve was completely transected below the ligation site with fine surgical scissors. Rats were anesthetized with an intraperitoneal injection of the ketamine and xylazine mixture (90 mg/kg and 8mg/kg, respectively). SNI in rats was performed following the same procedures as for mice. To create a dorsal column lesion in the spinal cord, a dorsal laminectomy was performed at the T9 level and bilateral dorsal columns with adjacent lateral columns were cut out with iridectomy scissors. To visualize regenerating axons, cholera toxin subunit B (CTB; List Biological Laboratories) was injected using a protocol modified slightly from that in the previous report 21. Briefly, after the sciatic nerve between the thigh muscles was exposed, a small incision was made on the perineurium just proximal to the trifurcation site. Two microliters of unconjugated CTB solution (1% in PBS) were slowly injected using the Hamilton syringe through the perineural incision and animals were killed 5 d after the injection.

• Product #104 – Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt